Influenza virus is an RNA virus belonging to the family Orthomyxoviridae. The viral RNA consists of eight independent segments, which easily recombine among influenza viruses to produce new subtypes.
Nucleoprotein (NP), which is the primary component of the nucleocapsid, is encoded in the fifth segment. The NP and the matrix protein are used to classify the influenza virus into group A, B or C. Since NP is an internal protein, it is not subject to the pressure of selection by a host's immune system. It binds RNA, is part of the transcriptase complex, and is involved in the nuclear-cytoplasmic transport of viral RNA (vRNA).
Neuraminidase (NM), which splits the α-keto bond that joins a terminal sialic acid and the next sugar residue, thereby allowing the release of viral progeny from infected cells, is encoded by the sixth segment. Nine subtypes (N1-N9) of this enzyme have been identified. All subtypes have two structural regions—a stalk and a head. All N8 proteins have 470 amino acids, the first eight of which are highly conserved. The following region is rich in hydrophobic amino acids and is considered to be the transmembrane domain. The next 51 amino acids make up the stalk region, and the head region begins at Cys91. The last region contains the catalytic site of the enzyme. Cysteine residues in the head and stalk region tend to be highly conserved. There are 6-8 putative N-glycosylation sites.
Hemagglutinin (HA), which is a membrane glycoprotein responsible for the adsorption of the virus into the host cell, is the main antigen to which neutralizing antibodies are directed. Its antigenic variation is the major cause of influenza epidemics. It is encoded by the fourth segment. Sixteen different subtypes (H1-H16) have been identified. HA has a signal peptide of 16 amino acids and two polypeptides (HA1 and HA2) joined by disulfide bridges. HA1 has the amino terminal end, whereas HA2 has the carboxyl terminal end. A hydrophobic region in HA2 anchors HA to the viral membrane. Cysteine residues tend to be highly conserved. There are six putative glycosylation sites, which enable the virus to mask its antigenic sites (Skehel et al., PNAS USA 81: 1779 (1984)).
Other proteins include matrix (M or M1 and M2), non-structural (NS or NS1 and NS2), PA, PB1, and PB2. The M1 protein is a major component of the virion that binds to the plasma membrane of infected cells by means of two hydrophobic regions at the N-terminus of the protein, whereas M2 is an ion channel and, therefore, an integral membrane protein. The NS1 protein is found in the nucleus and affects cellular RNA transport, splicing, and translation. The NS2 protein is found in the nucleus and cytoplasm and has unknown function. The PA protein is a transcriptase and may have protease activity, whereas the PB1 protein functions in transcription elongation and the PB2 protein functions in transcription cap binding.
Globally, influenza is the most economically significant respiratory disease in humans, pigs, horses and poultry (Wright et al., Orthomyxoviruses. In: Fields Virology. Knipe et al., eds. Lippincott Williams & Wilkins, Philadelphia, 2001. pp. 1533-1579.). Influenza virus is known for its continuous genetic and antigenic changes, which impede effective control of the virus (Wright et al. (2001), supra; Webster et al., Microbiol. Rev. 56: 152-179 (1992)). Of particular concern for prevention of epidemics and pandemics is the emergency of a new subtype of the virus by genetic re-assortment or inter-species transmission (Wright et al. (2001), supra).
Recently, influenza outbreaks have occurred in species, e.g., feline and canine, which historically do not carry influenza virus (Keawcharoen et al., Emerg. Infect. Dis. 10: 2189-2191 (2004); Crawford et al., Science 310: 398-485 (Oct. 21, 2005; published online Sep. 29, 2005); Dubovi et al., Isolation of equine influenza virus from racing greyhounds with fatal hemorrhagic pneumonia. In: Proceedings of the 47th Annual Meeting of American Association of Veterinary Laboratory Diagnosticians, Greensboro, N.C., Oct. 2005. p. 158; and Yoon et al., Emerg. Infect. Dis. 11(12): 1974-1976 (Dec. 2005)). Therefore, the host range of influenza virus is expanding.
Outbreaks of respiratory disease in racing greyhounds caused by infection with influenza virus have occurred in Florida in 2004, in eastern and western Iowa in April 2005, and in Texas in 2005. The disease was characterized by rapid onset of fever and cough, rapid respiration, and hemorrhagic nasal discharge. The morbidity was almost 100% in both race track compounds in Iowa, although the mortality was less than 5%. While a large percentage of affected dogs recovered, many succumbed to hemorrhagic pneumonia. Therapeutic administration of broad-spectrum antibiotics reduced the severity of the disease but could not control it.
In view of the above, it is an object of the present invention to provide the influenza virus that infects canines. It is another object of the present invention to provide materials and methods for inducing an immune response to the influenza virus in canines. These and other objects and advantages, as well as additional inventive features, will become apparent from the detailed description provided herein.